To initiate disease, the pathogenic Neisseria must invade non-ciliated cells of the mucosal epithelium, transcytose across the cell and exit into the subepithelial stroma. Cell culture and human volunteer studies demonstrate the N. gonorrhoeae have extended contact with its host, on the mucosal surface, and within cells. The Principal Investigator has been studying an important aspect of the intracellular life cycle of Neisseria: how the bacteria avoid lysosome killing. Their experiments demonstrate that the secreted Neisserial type 2 IgA1 protease is important for intracellular survival, as an iga mutant fails to replicate intracellularly. The protease also cleaves LAMP1 at its IgA1-like hinge, thereby accelerating its degradation and reducing its steady state levels in infected cells. LAMP1 is an integral lysosomal membrane glycoprotein which has been hypothesized to play a role in protecting the lysosomal membrane from degradation by its resident acid hydrolases. The Principal Investigator's results are consistent with this hypothesis. They indicate that infection results in multiple changes to lysosomes, as judged by decreased levels of several lysosomal constituents. they also show that the IgA1 protease is indirectly involved in these reductions. In this proposal, the Principal Investigator will continue her studies on Neisseria intracellular survival. She will determine how IgA1 protease reaches LAMP1 compartments, determine whether other secreted Neisserial proteases are capable of hydrolyzing LAMP1, identify the LAMP1 cleavage sites for the IgA1 protease, determine whether the protease is important for transcytosis and determine whether Neisseria escape the phagosome. her studies should shed light on the function of the Neisserial protease(s) in intracellular survival. They may also help to determine whether the IgA1 protease is a worthwhile target for a gonococcal vaccine. As many other bacterial pathogens secrete proteases of unknown function with similar specificities for human IgA1, her studies may shed light on the function of a large class of bacterial proteases.